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Agrobacterium-mediated transformation of a Eucalyptus camaldulensis × E. tereticornis hybrid using peeled nodal-stem segments with yeast HAL2 for improving salt tolerance

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Abstract

Genetic improvements of Eucalyptus require both breeding and gene transformation to generate desirable traits for forest industry. Because most of the current Eucalyptus transformation methods rely on juvenile seedling tissues as explants, the resulting transgenics are subjected to phenotypic segregation through the use of segregated seedlings. This is unsuitable for transformation of selected Eucalyptus breeding lines, especially the interspecific hybrids. Here, we developed an Agrobacterium-mediated transformation method using peeled nodal-stem explants from in vitro-propagated plantlets of a Eucalyptus hybrid. The explant was prepared by peeling off epidermal layers around the axillary buds using a pair of forceps, before co-cultivation with Agrobacterium tumefaciens. Transgenic shoots generated from the axillary buds were then obtained through kanamycin selection. The transformation efficiency was at most 24% as confirmed by PCR and GUS assay. Transformation of a E. camaldulensis × E. tereticornis F1 hybrid clone using yeast HAL2 showed that, out of 35, 11 transgenic plantlets had an increase in salt tolerance and grew and developed roots, though at a slow rate, on the medium supplemented with NaCl up to 300 mM concentration. This work demonstrates the efficiency of this transformation method for further genetic improvement of Eucalyptus clones selected through hybridization and breeding.

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Acknowledgements

This research was supported by The Graduate School for Thesis and Dissertation, Kasetsart University, Kasetsart University Research and Development Institute (KURDI), and Faculty of Science Research Fund (ScRF).

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Authors and Affiliations

  1. Faculty of Science and Technology, Valaya Alongkorn Rajabhat University under the Royal Patronage, 20 Phaholyothin Road, Klong Nuang, Khlong Luang, Pathum Thani, 13180, Thailand

    Narumol Thanananta

  2. Department of Genetics, Faculty of Science, Kasetsart University, 50 Ngam Wong Wan Road, Chatuchak, Bangkok, 10900, Thailand

    Supachai Vuttipongchaikij & Somsak Apisitwanich

  3. Center of Advanced Studies for Tropical Natural Resources, Kasetsart University, 50 Ngam Wong Wan Road, Chatuchak, Bangkok, 10900, Thailand

    Supachai Vuttipongchaikij & Somsak Apisitwanich

  4. School of Science, Mae Fah Luang University, Chiangrai, 57100, Thailand

    Somsak Apisitwanich

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  1. Narumol Thanananta
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  2. Supachai Vuttipongchaikij
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  3. Somsak Apisitwanich
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Correspondence to Supachai Vuttipongchaikij or Somsak Apisitwanich.

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11056_2017_9621_MOESM1_ESM.tif

Supplementary data Fig. S1 shows vigorous growth of the 24A17 hybrid compared with its parental lines, E. calmaldulensis and E. tereticornis, and other two F1 hybrid lines (TIFF 10966 kb)

11056_2017_9621_MOESM2_ESM.tif

Supplementary data Fig. S2 shows growth of the 24A17 hybrid, E. calmaldulensis and E. tereticornis cultivated on the growth medium supplemented with 0, 100, 300 and 500 mM NaCl for 30 days (TIFF 9342 kb)

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Thanananta, N., Vuttipongchaikij, S. & Apisitwanich, S. Agrobacterium-mediated transformation of a Eucalyptus camaldulensis × E. tereticornis hybrid using peeled nodal-stem segments with yeast HAL2 for improving salt tolerance. New Forests 49, 311–327 (2018). https://doi.org/10.1007/s11056-017-9621-5

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